Liquid chromatography column

ABSTRACT

A chromatography cartridge assembly includes a cartridge and first and second end caps. A wall of the cartridge defines a chamber for containing chromatography media. The end caps define inlet and outlet passages, respectively, for flow of process fluid. The first end cap defines flow distributor passages for distributing process fluid across a cross-sectional area of the chamber, and the second end cap defines flow collector passages for collecting process fluid from across a cross-sectional area of the chamber. A clamp is circumferentially located about the cartridge wall for applying a radial load through the wall to the first end cap to fix the first end cap in a desired position.

This application is a continuation-in-part of application U.S. Ser. No.08/970,287, filed Nov. 14, 1997 now abandoned, and application, U.S.Ser. No. 08/970,286, filed Nov. 14, 1997, now abandoned.

BACKGROUND OF THE INVENTION

This invention relates to liquid chromatography columns.

Cartridges for use in liquid chromatography are known in which a flowdistributor and a flow collector are located in the vicinity of the endcaps of the cartridge to distribute and collect the process fluid.

SUMMARY OF THE INVENTION

The invention features, in general, a chromatography cartridge assemblyincluding a cartridge and a first end cap. A wall of the cartridgedefines a chamber for containing chromatography media. The first end capis positioned within a first end cap receiving opening of the cartridge.The first end cap defining a passage for flow of process fluid. A clampis circumferentially located about the cartridge wall for applying aradial load through the wall to the end cap to fix the end cap in adesired position.

In preferred embodiments, a second end cap is positioned within a secondend cap receiving opening of the cartridge. The second end cap defines apassage for flow of process fluid. A clamp is circumferentially locatedabout the cartridge wall for applying a radial load to the second endcap to fix the second end cap in a desired position.

The first end cap includes flow distributor passages for distributingprocess fluid across a cross-sectional area of the chamber, and thesecond end cap includes flow collector passages for collecting processfluid from across a cross-sectional area of the chamber.

The inner surface of the wall has a constant diameter. Alternatively,the inner surface of the wall defines a section of constant innerdiameter for slidably receiving the first end cap, and a circumferentialgroove for receiving the second end cap. Alternatively, the secondunitary end cap is integral with the cartridge.

The end caps each include a sieve for retaining the chromatography mediain the chamber. The sieve includes a fine mesh and a course mesh.

Flexible seals are located between the end caps and the wall of thecartridge. The first and second end caps each include a connector forattachment to an inlet conduit and an outlet conduit, respectively.

A support clamp holds the cartridge. The cartridge includes a flexiblewall configured to radially compress the chromatography media. Thechromatography media is a hydrophilic material. The chromatography mediahas an operating pressure rating greater than about 3 bar. Thechromatography media has a particle size in the range of about 15-200microns.

According to another aspect of the invention, a chromatography apparatusincludes a cartridge assembly and a compression module surrounding thecartridge assembly. The compression module defines a pressure chamberfor containing a pressurized fluid. The pressurized fluid acts to move aflexible wall of the cartridge.

According to another aspect of the invention, a method of revitalizing apacked column having trapped air includes providing a chromatographycartridge including a flexible wall defining a chamber. The flexiblewall forms a movable diaphragm for compressing chromatography medialocated within the chamber. Compression is applied to the chromatographymedia to minimize the volume of trapped air.

In preferred embodiments, the compression is radial compression.

According to another aspect of the invention, a method of sealing aflexible-walled tube includes placing a sealing member within the tube,positioning a first conical member around the outside of the tube andaxially aligned with the sealing member, and positioning a secondconical member around the outside of the tube and in contact with thefirst conical member. An axial load is applied to the second conicalmember, whereby the axial load on the second conical member acts toradially compress the first conical member. The radial compressioncauses permanent deformation of the first conical member and theadjacent wall of the tube to seal the sealing member within the tube.The seal is maintained upon removal of the axial load.

In preferred embodiments, the sealing member is an end cap defining aninlet passage for flow of process fluid and flow distributor passagesfor distributing process fluid across a cross-sectional area of thetube.

According to another aspect of the invention, a clamping mechanism forsealing a sealing member within a tube includes a first conical memberfor placement around the outside of the tube, a second conical memberfor placement around the outside of the tube and in contact with thefirst conical member, and a load applicator for applying an axial loadto the second conical member. The axial load on the second conicalmember acts to radially compress the first conical member. The radialcompression causes permanent deformation of the first conical member andthe adjacent wall of the tube to seal the sealing member within thetube.

According to another aspect of the invention, a chromatography columnincludes a column having a flexible-wall. The column containschromatography media. An end cap is positioned within the column anddefining an inlet passage for flow of process fluid and flow distributorpassages for distributing process fluid across a cross-sectional area ofthe column. A clamp is located around an outside of the column forsealing the end cap within the column.

In preferred embodiments, the clamp radially compresses the flexiblewall to seal the end cap within the column. The clamp is a conicalmember.

Advantages include a cartridge which can be dynamically compressed andused as a stand alone device. The cartridge is disposable and providesconvenience over glass columns which the user packs themselves. Thecartridge seals are static and thus easier to clean between runs thanthe dynamic seals in a glass column.

Additional advantages include liquid chromatography of biomoleculeprocess fluids under pressures above 3 bar. The materials used in thechromatography apparatus prevent biomolecule precipitation andnon-specific adsorption. The cartridge within a module system permitschangeout of wetted components and reuse of the module for differentbiomolecules without cross-contamination.

Other advantages and features of the invention will ba apparent from thefollowing description of a preferred embodiment thereof and from theclaims.

BRIEF DESCRIPTION OF THE DRAWINGS

The drawings will be described first.

Drawings

FIG. 1 is a schematic of a chromatography apparatus according to theinvention;

FIG. 2 is a partially cut-away, cross-sectional side view of thepressure module of the invention;

FIG. 2A is a top view of the pressure module of FIG. 2;

FIG. 2B is an enlarged view of the clamping region of the pressuremodule of FIG. 2;

FIG. 3 is a partially cut-away, cross-sectional side view of a cartridgeassembly of the invention;

FIG. 4 is a cross-sectional side view of a distributor and mesh of theinvention;

FIG. 4A is a sectional view of the distributor of FIG. 4, taken alonglines 4A—4A;

FIG. 5 is an enlarged schematic view of a sealing scheme of theinvention; and

FIG. 6 is an enlarged schematic view of an additional sealing scheme ofthe invention.

FIG. 7 is shows an additional embodiment of a chromatography assemblyaccording to the invention;

FIG. 8 is a cross-sectional side view of the cartridge and unitary endcaps of the chromatography assembly of FIG. 7;

FIG. 9 is a cross-sectional, exploded side view of a unitary end cap ofthe chromatography assembly of FIG. 7;

FIG. 10 is a top view of a support clamp of the chromatography assemblyof FIG. 7;

FIG. 10A is a cross-sectional side view of the clamp of FIG. 10, takenalong lines 10A—10A;

FIG. 11 is a cross-sectional side view of a section of a pressure modulemodified for use with chromatography assembly of FIG. 7;

FIG. 12 is a cross-sectional side view of an additional embodiment of achromatography cartridge;

FIGS. 13 a and 13 b are cross-sectional side views of the chromatographycartridge of FIG. 12 mounted to a stand;

FIGS. 14 a and 14 b are side and top views, respectively, of a loadapplicator for clamping an end cap of the chromatography cartridge ofFIG. 12; and

FIGS. 15 and 16 show various stages in the clamping process.

DESCRIPTION OF THE PREFERRED EMBODIMENT

Referring to FIG. 1, an apparatus 10 is shown for performingchromatography separation of biomolecules, e.g., proteins,oligosaccharides, large DNA molecules, and viral particles, in anaqueous based solvent. The term biomolecules is not meant to includesynthetic organic chemicals, small linear peptides, or chiral compounds.Apparatus 10 includes a chromatography assembly 12 and inlet solutiontank 2, load tank 3, and system pump 4 for delivering process fluidunder pressure along a process inlet path 14 to chromatography assembly12. An outlet line 5 leads from chromatography assembly 12 to a productcollection vessel 6 and a waste receptacle 7. A water filter, bubbletrap and monitor 8 (monitoring, e.g., pressure, conductivity, and pH)are located along the process fluid inlet path 14. A monitor 9monitoring, e.g., pressure, conductivity, pH, and UV absorbance, islocated along outlet line 5. A column bypass 16 permits the system to becleaned while bypassing the chromatography assembly. Valves 15 controlthe flow of the process fluid.

Referring to FIGS. 2-2B, chromatography assembly 12 includes acompression module 20 and a cartridge assembly 22. Compression module 20includes a housing 30, formed from, e.g., stainless steel or aluminum,defining a cylindrical region 32 for containing fluid for applyingradial compression to cartridge assembly 22. A compressible orincompressible fluid can be used to apply radial compression pressure tocartridge assembly 22.

The application of radial compression to a chromatography cartridge isdescribed in U.S. Pat. No. 4,250,035 to McDonald, hereby incorporated byreference. Briefly, in a liquid chromatography column, a stationaryphase such as silica is packed in a cartridge having a flexible wall. Byexerting radial pressure on the cartridge, packing bed voids are avoidedand wall channeling effects are overcome. The packing efficiency of thecolumn is increased and is more reproducible, and greater uniformity canbe achieved in column performance both among packed columns of the samekind and during the useful life of a given packed column.

Referring to FIGS. 1 and 2A, housing 30 includes a fluid inlet 34, arelief valve 36 for purging pressure within cylindrical region 32, and apressure indicator 38. Radial compression pressure applied to cartridgeassembly 22 is controlled by a pressure regulator or a pump (not shown)which delivers fluid to fluid inlet 34; solvent flow rate through thecartridge assembly is controlled by pump 4. A mounting arm 74 connectedto housing 30 can be used to mount chromatography assembly 12 to alaboratory stand.

Removable end caps 40, 42 retain cartridge assembly 22 in place withincompression module 20. Referring particularly to FIG. 2B, end cap 42 ismounted to housing 30 with a band clamp 44 (end cap 40 is similarlymounted to housing 30 with a band clamp 46). Clamp tightening knobs 48,50 are used to tighten band clamps 44, 46 respectively. At higherpressures, the knobs can be replaced with bolts to meet coderequirements. As shown in FIG. 2B, each end cap 40, 42 is sealed againsthousing 30 with an o-ring 52 to prevent leakage of compression fluidfrom region 32. As shown in FIG. 2, end caps 40, 42 are sealed againstcartridge assembly 22 by o-rings 53, 55, respectively, which separatecompression fluid from process fluid.

An inlet connector 60 defines a channel 61 leading to an inlet passage62 defined by end cap 40 for flow of process fluid into cartridgeassembly 22. Control knob 64 is used to open and close channel 61. Anoutlet passage 66 defined by end cap 42 leads to an outlet connector 68defining a channel 69 for flow of process fluid out of cartridgeassembly 22. Inlet and outlet passages 62 and 66 include o-ring seals70, 72, respectively, for sealing the passages against cartridgeassembly 22. End caps 40, 42 are preferably made from a hydrophilicmaterial, e.g., stainless steel, to prevent precipitation ofbiomolecules on the surfaces of passages 62, 66. Seals 70 and 72 preventflow of process fluid along the interface 73 between end cap 40 andcartridge 22 and the interface 75 between end cap 42 and cartridge 22thus minimizing the exposure of the process fluid to dead spaces andcrevices in which microbial growth and attachment could occur.

Referring to FIG. 3, cartridge assembly 22 has a flexible wall 80partially defining a media chamber 82. Flexible wall 80 further definesend cap receiving openings 84, 86. The upper and lower ends 88, 90 ofmedia chamber 82 are defined by flow assemblies 92, 94 respectively.Upper flow assembly 92 includes a flow distributor 100 and a sieve 102,e.g., a mesh or frit. A mesh is preferred over a frit due to its smallersurface area which limits biomolecule adhesion. Lower flow assembly 94includes a flow collector 104 and a sieve 106. The flow distributor 100,flow collector 104, and sieves 102, 106 are preferably made fromhydrophilic materials having surface energies greater than about 36dyn/cm, e.g., polyamide, polyethyleneterephthalate, polyvinylidenechloride, polymethylmethacrylate, and polystyrene, to limit biomoleculebinding to the surfaces and clogging of the sieves. Materials havingsurface energies less than 36 dyn/cm are not suitable for separatingbiomolecules because the biomolecules adhere to the material thusclogging the cartridge assembly.

Referring to FIGS. 4 and 4 a, sieve 102 is welded to flow distributor100 along outer periphery 103 of flow distributor 100. Welding alongperiphery 103 permits process fluid to flow through sieve 102 but notaround it, and prevents media particles from leaking around sieve 102into flow distributor 100. Sieve 106 is similarly welded to flowcollector 104.

The process fluid path is from inlet passage 62 to an inlet 110 of flowdistributor 100. Multiple flow channels 112, 8 channels being shown inthe illustrated embodiment, run from inlet 110 to outlets 114. Outlets114 connect flow channels 112 to a network of channels 116 whichdistribute the process fluid. Sieve 102 preferably has a pore size ofabout 10-20 micron to allow passage of process fluid while preventingpassage of chromatography media. Flow collector 104 and sieve 106 areidentical to flow distributor 100 and sieve 102. Flow collector 104 andsieve 106 are mounted such that process fluid first passes through sieve106 and then through the network of channels 116 to finally be collectedat inlet 110.

An alternative or additional sealing scheme which further limits voidsand dead spaces in which process fluid can be trapped is shown in FIG.5. Here, an o-ring 121 positioned between flow distributor 100 andcartridge wall 80 prevents flow of process fluid around edge 123 of theflow distributor and into crevices where the process fluid can betrapped. Similarly, an o-ring can be positioned between flow collector104 and cartridge wall 80. Additionally, referring to FIG. 6, the flowdistributor and/or flow collector can be welded at 130 along edge 123and side portions 132, 134 to the cartridge wall, thereby creating a lowdead volume seal.

Example operating pressure (process fluid flow pressure) ratingsachievable with chromatography assembly 12 employing an aluminumcompression module 20 are listed below. For an incompressiblecompression fluid, the operating pressure can be equal to the pressurerating of the pressure module. For a compressible compression fluid, theoperating pressure is about 1 to 6 bar less than the pressure rating ofthe pressure module because the compression pressure applied to thecartridge is greater than the process fluid pressure to maintain theintegrity of the cartridge. Higher pressure ratings are achievabledepending upon tube thickness and by substituting stainless steel foraluminum.

inner diameter of compression module 20 (mm) pressure (bar)  75 20-35100 14-23 150 10-17 300  6-14 400  4-10

Referring again to FIG. 3, chromatography media 120 is contained withinmedia chamber 82 by upper and lower sieves 102, 106. Due to recentadvances in materials technology leading to the development of the newhydrophilic and rigid support matrices having high pressure ratings, thehigh pressure ratings achievable with chromatography assembly 12 and thehydrophilic materials used in the critical components of cartridgeassembly 22 enable fast, high resolution biomolecule separation.Suitable matrices for chromatography media 120 include Emphaze™,available from Pierce; POROS®, available from PerSeptive Biosystems;HyperD™, available from BioSepra; Source™, available from PharmaciaBiotech, Sweden; Toyopearl®, available from TosoHaas; Fractogel®,available from E. Merck, Germany, Macro-Prep®, available from BioRad;Bakerbond®, available from Baker Mallinckrodt; Sepharose®, availablefrom Pharmacia Biotech; and Amberchrom, available from TosoHaas. Themedia listed above have particle sizes in the range of 15-100 microns,though media can be used having larger particle sizes, up to at leastabout 200 microns. Another suitable matrix for chromatography media 120is a continuous bed matrix, e.g., the UNO Continuous Bed Matrix,available from BioRad. The pressure ratings and availablefunctionalities of each material are listed below.

Pressure Available Matrix Rating (bar) Functionalities POROS ® 100 ionexchange hydrophobic interaction affinity HyperD ™ 200 ion exchanqeaffinity Emphaze ™ 7 affinity Fractogel ® 10 ion exchange hydrophobicinteraction affinity Toyopearl ® at least 7 ion exchange hydrophobicinteraction affinity Source ™ 50 ion exchange hydrophobic interactionMacro- 55 ion exchange Prep ® Bakerbond ® 70-140 ion exchangehydrophobic interaction Sepharose ® 3 ion exchange hydrophobicinteraction affinity Amberchrom Not Available reverse phase UNO Matrix48 ion exchange

Sepharose has a pressure rating of about 3 bar. All the other media havepressure ratings above 3 bar, above 5 bar, and some have pressureratings about 50 bar, with one having a rating greater than 150 bar.

The radial pressure applied to the chromatography media should be atleast equal to the flow pressure of the process fluid to maintain theintegrity of the column. When using a compressible compression fluid,the radial pressure applied is in the range of about 1 to 6 bar over theoperating pressure.

The surfaces of chromatography assembly 12 exposed to process fluidinclude cartridge 22, flow distributor 100, flow collector 104, sieves102, 106, and end caps 40, 42. As discussed previously, the flowdistributor, flow collector and sieves are formed from hydrophilicmaterials to prevent biomolecule precipitation and non-specificadsorption. The sieves are preferably polymeric as opposed to stainlesssteel due to the stainless steel's poorer chemical resistance andsusceptibility to chloride attack. Because the surface area of cartridge22 exposed to the process fluid is much less than that of the flowdistributor, flow collector, and sieves, cartridge 22 can be formed froma less hydrophilic material, e.g., polyethylene having a surface energyof 35.7 dyn/cm (linear PE) and 35.3 dyn/cm (branched PE). Though tominimize biomolecule precipitation and non-specific adsorption on thecartridge, preferably a more hydrophilic material is also used forcartridge 22. End caps 40, 42 are preferably stainless steel.

Seals 53, 55, 70, 72 insure that compression module 20 remains free ofcontamination from process fluid during use. The components of cartridgeassembly 12 with wetted surfaces can be changed while the samecompression module 20 can be used with a new sample withoutcross-contamination.

It is understood that separate inserts can be employed to definepassages 62, 66 such that end caps 40, 42 are not exposed to processfluid and only the inserts need be removed and exchanged or cleanedbetween sample runs.

It has been found that radial compression can revitalize a packedcolumn. Trapped air in the media causes bed cracking and loss ofchromatographic efficiency. By subjecting the column to radialcompression, the volume of air is minimized thus minimizing the effectsof air entrapment such that there is little or no decrease inperformance of the column.

In another embodiment of the invention, referring to FIGS. 7 and 8, achromatography assembly 212 includes a cartridge 222 and unitary endcaps 240, 242. Cartridge 222 has a flexible wall 280 partially defininga media chamber 282. Flexible wall 280 further defines end cap receivingopenings 284, 286. Cartridge 222 can be used alone or with a compressionmodule, as described above. When used alone, cartridge 222 is supportedby one or more mechanical support clamps 220. Clamps 220 can beconnected to a stand 274, as described further below.

Cartridge assembly 212 can be formed of hydrophilic materials havingsurface energies greater than about 36 dyn/cm when used for separatingbiomolecules, as described above, and may be formed of other materials,e.g., polyethylene or stainless steel, when denaturation of biomoleculesis not a concern.

Upper unitary end cap 240 includes an end cap body 241 defining a flowdistributor section 300. Flow distributor section 300 is similar to flowdistributor 100, described above with reference to FIGS. 4 and 4A, butin this embodiment the flow distributor is formed directly in the endcap body 241. A sieve 302 is welded to end cap body 241 along outerperiphery 303 of end cap body 241 (see FIG. 9). Lower unitary end cap242 similarly includes an integral flow collector 304 and a sieve 306.

Referring to FIG. 9, sieve 302 preferably includes a fine mesh 307having a pore size of about 5-10 micron to allow passage of processfluid while preventing passage of chromatography media, and a coarsemesh 309 having a pore size of about 800 micron and provides support forthe fine mesh. Flow collector 304 and sieve 306 are identical to flowdistributor 300 and sieve 302.

Referring again to FIG. 8, cartridge wall 280 defines a first section380 having a constant inner diameter, e.g., 3.187″, and a second section382 of decreased inner diameter, e.g., 3.01″, resulting in an increasein the thickness of cartridge wall 280. Lower unitary end cap 242 ismechanically pressed into a circumferential channel 384, e.g., 0.11″deep, in wall 280. Alternatively, lower unitary end cap 242 can beformed integrally with cartridge wall 280, e.g., by molding the elementsas a single unit, eliminating the need for o-ring 255, described below.The cartridge wall is dimensioned to provide flexibility so that thecartridge can be radially compressed and to provide a rigid wall whenthe cartridge is under ambient conditions. Cartridge wall 280 hasdimensions of, e.g., a thickness t₁ of 0.1565″, t₂ of 0.2065″, t₃ of0.295″, t₄ of 0.25″, and t₅ of 0.1625″. Cartridge 222 has an outerdiameter D₁ of, e.g., 3.60″.

Each unitary end cap 240, 242 is sealed against wall 280 of cartridge222 with an o-ring 253, 255, respectively to prevent leakage of processfluid between wall 280 and the end caps. Upper unitary end cap 240defines a circumferential groove 270 in which o-ring 253 is located.O-ring 255 is located between wall 385 of channel 384 and acircumferential chamfer 272 defined by lower unitary end cap 242.

Upper unitary end cap 240 is slidably received within section 380 ofcartridge 222 and is fixedly mounted to cartridge 222 with a band clamp244, e.g., a blade draw latch type clamp or a J-style preformed hoseclamp available from McMaster-Carr.

Upper unitary end cap 240 defines an inlet connector 260 and an inletpassage 262 for flow of process fluid into cartridge 222. Lower unitaryend cap 242 defines an outlet connector 268 and an outlet passage 266for flow of process fluid out of cartridge 222. The unitary end capdesign minimizes the exposure of the process fluid to dead spaces andcrevices in which microbial growth and attachment could occur.

Referring to FIGS. 10 and 10A, clamp 220 has an inner diameter D₂ of,e.g., 3.650″, and a space S₁ of e.g., 0.150″ such that it holdscartridge 222. A mount 221 of clamp 220 permits the attachment of theclamp to support 274 as well as to any variety of supports to stabilizethe cartridge during use and shipping. Clamp 220 can be secured aboutcartridge 222 with a closure (not shown) such as a screw or a singlelatch-type buckle.

Referring to FIG. 11, compression module 20 of FIG. 2 is shown modifiedto accommodate the unitary end caps of the cartridge assembly of FIG. 7.Removable end cap 40 of FIG. 2 have been replaced with a removablemodule sealing adapter 340 and a removable module end cap 341 (removableend cap 42 of FIG. 2 is similarly replaced with an identical removablemodule sealing adapter and end cap, not shown).

Adapter 340 and end cap 341 are mounted to module housing 330 with aband clamp 344. Sealing adapter 340 is sealed against unitary end cap240, module end cap 341 is sealed against adapter 340, and module endcap 341 is sealed against housing 330, by o-rings 350, 351 and 352,respectively. Sealing adapter 340 and end cap 341 include matingthreaded sections 360, 362 with o-ring 351 providing a sliding sealbetween sealing adapter 340 and end cap 341. The threaded connectionbetween sealing adapter 340 and end cap 341 provides the degree offreedom necessary to adjust their relative positions dependent upon thefinal position of upper unitary end cap 240. Sealing adapter 340 andmodule end cap 341 are preferably formed from stainless steel.

Prior to filling cartridge 222 with chromatography media, lower unitaryend cap 242 is positioned in cartridge 222. Cartridge 222 is then filledwith chromatography media to a desired column height. Column packing maybe performed as described in commonly owned U.S. patent application,U.S. Ser. No. 08/970,286, entitled CHROMATOGRAPHY MEDIA PACKING SYSTEM,filed Nov. 14, 1997, which is incorporated herein by reference. Upperunitary end cap 240 is then slid into cartridge 222 to rest against thechromatography media. The performance characteristics of the packedcartridge are then tested. If the column does not perform as desired,upper unitary end cap 240 can be removed and the column repacked. Afterthe desired performance characteristics are obtained, upper unitary endcap 240 is held in place with band clamp 244.

An alternative method of fixing the end caps in place will now bedescribed. Referring to FIG. 12, a chromatography assembly 412 includesa cartridge 422 and sealing members, e.g., unitary end caps 440, 442,fixed to cartridge 422 with clamps 444, 446, respectively. Unitary endcaps 440, 442 are similar to unitary end caps 240, 242, described above,having an integral flow distributor and sieve (e.g., a mesh or frit),and integral flow collector and sieve (e.g., a mesh or frit),respectively.

Cartridge 422 has a flexible wall 480 with a constant inner diameter inthe range of, e.g., about 0.472″ to 16″, an outer diameter in the rangeof, e.g., about 0.63″ to 16.375″, and a wall thickness in the range of,e.g., about 0.08 to 0.375″. Cartridge 422 can be made from, e.g.,polyproplyene, polyetheylene, nylon, or thin-walled (e.g., 0.03″)stainless steel.

Unitary end caps 440, 442 are slidably received within cartridge 422 andfixed at desired locations within cartridge 422 with clamps 444, 446,respectively, as described below. Each unitary end cap 440, 442 issealed against wall 480 of cartridge 422 with an o-ring 453, 455,respectively, to prevent leakage of process fluid between wall 480 andthe end caps. Alternatively, lower unitary end cap 442 can be formedintegrally with cartridge wall 480, e.g., by molding the elements as asingle unit.

Prior to filling cartridge 422 with chromatography media, lower unitaryend cap 442 is positioned in cartridge 422 and clamped in place withclamp 446, as described below. Cartridge 422 is then filled withchromatography media to a desired column height. The column is thenpacked, with upper unitary end cap 440 being slid into cartridge 422 torest against the chromatography media. After the desired performancecharacteristics are obtained, upper unitary end cap 440 is fixed inplace with clamp 444.

Referring to FIGS. 13 a and 13 b, lower clamp 446 is shown in positionfixing lower end cap 442 within cartridge 422, and upper clamp 444 isshown prior to clamping of upper end cap 440. Clamp 444 (as well asclamp 446) is in the form of a ring 520 having a cylindrical inner wall522 and a conical outer wall 524. To fix end cap 440 in place, ring 520is slid over cartridge wall 480 and positioned in the vicinity of o-ring453. An axial load, in the direction of arrow, A, is then applied toring 520. Due to the conical shape of ring 520, the axial load isconverted to an inward, radial compressive load in ring 520. Thecompressive load acts to permanently deform ring 520 and portions ofcartridge wall 480 adjacent ring 520. This deformation of cartridge wall480 acts to fix end cap 440 within cartridge 422.

A backup ring 530 axially supports and positions ring 520. Backup ring530 presses against a larger end 526 of ring 520. Backup ring 530 isstronger than ring 520 to withstand the load applied to ring 520 withoutdeforming. Backup ring 530 preferable includes two or more segmentswhich permit backup ring 530 to be removed after use without the need toslide the backup ring to the end of cartridge 422, which may beobstructed by another compressed ring or other items.

Backup ring 530 is located within a counter-bore 542 of a support tube540. Support tube 540 has a wall 544 defining a bore 546 through whichcartridge 422 passes. An inner surface 548 of wall 544 is threaded at550. Support tube 540 is attached to a stand 552 by a support arm 560.Support arm 560 has a wall 564 defining a bore 566 through whichcartridge 422 passes. An outer surface 568 of wall 564 is threaded at570. Rotation of support tube 540 relative to support arm 560 permitsaxial adjustment of the position of backup ring 530 along cartridge 422.

To apply the axial load to ring 520, a compressing ring 580 having aninner conical wall 582 complementary to conical wall 524 of ring 520 ispositioned over ring 520 opposite backup ring 530. Compressing ring 580is stronger than ring 520 to withstand the load applied to ring 520without deforming. Compressing ring 580 can also be composed of two ormore segments, if necessary.

The angle of inner conical wall 582 of compressing ring 580 should matchthe angle of conical wall 524 of ring 520 if a flat compression againstwall 480 is desired. A 14 degree cone angle (28 degree included angle)has been found to be suitable. Larger angles require more axial force toapply the necessary compressive force. Smaller angles can make itdifficult to separate compressing ring 580 from ring 520 aftercompressing ring 520. A smaller angle also requires longer axial motionof compressing ring 580 to achieve the desired deformation of ring 520.The best angle for a given application depends on several factorsincluding the ring material (which should be malleable), surface finish,plating, lubrication, and available forces.

If an angled compression contact is desired, the angle of inner conicalwall 582 can be set at a different angle than that of conical wall 524.Alternatively, cylindrical wall 522 can be conical instead ofcylindrical to create an angled compression contact.

Referring to FIGS. 14 a and 14 b, a compressive force device 600 is usedto apply the axial load to compressing ring 580. Compressive forcedevice 600 includes a stationary frame 602 and a movable frame 603.Frame 602 includes four stationary, vertical struts 606, 608, 610 and612 and lower and upper stationary plates 614 and 616, respectively.Movable frame 603 is slidably mounted to frame 602. Movable frame 603includes two vertical plates 618, 620, lower horizontal struts 624, 626,and upper plate 628. A hydraulic cylinder 604 is mounted between lowerstationary plate 614 and lower movable struts 624, 626. When hydraulicfluid is pumped into cylinder 604, a piston rod 630 of cylinder 604extends. This acts to lift movable frame 603, forcing plate 628 closerto plate 616. A dial indicator 632 mounted to plate 628 measures thedistance between plate 628 and plate 616. Four springs 634 (two of thefour springs are shown in FIG. 14 a) act to lower movable frame 603 whenhydraulic pressure is removed. Plates 616 and 628 define cutouts 636which permit compressive force device 600 to be positioned aboutcartridge 422.

Referring again to FIG. 13 a, in use, backup ring 530 is positioned incounter-bore 542 of support tube 540. Ring 520 is slid over cartridgewall 480 and up against backup ring 530. Compressing ring 580 is thenside over cartridge wall 480 and onto ring 520. Compressing ring 580preferably has a thin film of high pressure grease on inner conical wall582. Support tube 540 is then rotated to move ring 520 until ring 520 isat the appropriate position relative to end cap 440. On clear ortranslucent flexible tubes, the ring may be positioned by sight. Onopaque tubes, an external indicator, not shown, can be used to positionthe ring, or the packing mechanism which positions the unitary end canbe used.

Referring to FIG. 15, compressive force device 600 is then positionedabout cartridge 422 and is supported by compressing ring 580. Hydraulicfluid is pumped into cylinder 604 to extend piston rod 630 (FIG. 16).This raises plate 628 toward plate 616, applying a load to ring 520.Hydraulic pressure is increased to about 1,000 psi to remove all slopand clearances from the system. Dial indicator 632 is then set to zero.Hydraulic pressure is increased until the desired deformation of ring520 is achieved, e.g., when the dial indicator reads 0.070″. The axialdeformation of 0.070″ corresponds to a radial compression of about0.034″.

Hydraulic pressure is then removed, and compressive force device 600,compressing ring 580, and cartridge 422 removed. The two halves ofbackup ring 530 will come off cartridge 422 when pulled out ofcounter-bore 542 of support tube 540.

The load required to deform ring 520 depends upon the size of ring 510.For example, for a ring 520 with an inner diameter of 0.645″, an outerdiameter which slopes from 0.725″ to 0.875″ at an angle of 14°, and alength of 0.3″, formed from 316 stainless steel, when the axial loadapplied to ring 520 reaches about 5300 pounds the yield point of thestainless steel is reached.

Ring 520 can be used in place of band clamp 244 of FIG. 8.

Other embodiments of the invention are within the scope of the followingclaims.

What is claimed is:
 1. A chromatography cartridge assembly comprising: acartridge including a wall defining a chamber for containingchromatography media, and a first end cap receiving opening, a first endcap received within said first end cap receiving opening, said first endcap defining a passage for flow of process fluid, and a clampcircumferentially located about said cartridge wall for applying aradial load through said wall to said end cap to fix said end cap in adesired position, said clamp including a conical member.
 2. Thechromatography cartridge assembly of claim 1 wherein said cartridgefurther defines a second end cap receiving opening, said assemblyfurther comprising a second end cap positioned within said second endcap receiving opening, said second end cap defining a passage for flowof process fluid.
 3. The chromatography cartridge assembly of claim 2further comprising a clamp circumferentially located about saidcartridge wall for applying a radial load to said second end cap to fixsaid second end cap in a desired position.
 4. The chromatographycartridge assembly of claim 2 wherein said second end cap includes asieve for retaining the chromatography media in said chamber.
 5. Thechromatography cartridge assembly of claim 4 wherein said second end capsieve comprises a fine mesh and a course mesh.
 6. The chromatographycartridge assembly of claim 2 further comprising a flexible seal locatedbetween said second end cap and said wall of said cartridge.
 7. Thechromatography cartridge assembly of claim 2 wherein said second end capincludes a connector for attachment to an outlet conduit.
 8. Thechromatography cartridge assembly of claim 1 wherein said first end capincludes flow distributor passages for distributing process fluid acrossa cross-sectional area of said chamber, and said second end cap includesflow collector passages for collecting process fluid from across across-sectional area of said chamber.
 9. The chromatography cartridgeassembly of claim 1 wherein an inner surface of said wall has a constantdiameter.
 10. The chromatography cartridge assembly of claim 1 whereinan inner surface of said wall defines a section of constant innerdiameter for slidably receiving said first end cap.
 11. Thechromatography cartridge assembly of claim 1 wherein said assemblyfurther comprises a second unitary end cap, said second end cap defininga passage for flow of process fluid, said second unitary end cap beingintegral with said cartridge.
 12. The chromatography cartridge assemblyof claim 1 wherein said first end cap includes a sieve for retaining thechromatography media in said chamber.
 13. The chromatography cartridgeassembly of claim 12 wherein said first end cap sieve comprises a finemesh and a course mesh.
 14. The chromatography cartridge assembly ofclaim 1 further comprising a flexible seal located between said firstend cap and said wall of said cartridge.
 15. The chromatographycartridge assembly of claim 1 wherein said first end cap includes aconnector for attachment to an inlet conduit.
 16. The chromatographycartridge assembly of claim 1 wherein said cartridge includes a flexiblewall.
 17. The chromatography cartridge assembly of claim 16 wherein saidflexible wall is configured to radially compress said chromatographymedia.
 18. The chromatography cartridge assembly of claim 1 furtherincluding chromatography media contained within said chamber.
 19. Thechromatography cartridge assembly of claim 18 wherein saidchromatography media comprises a hydrophilic material.
 20. Thechromatography cartridge assembly of claim 18 wherein saidchromatography media has an operating pressure rating greater than about3 bar.
 21. The chromatography cartridge assembly of claim 18 whereinsaid chromatography media has a particle size in the range of about15-200 microns.
 22. The chromatography cartridge assembly of claim 1wherein said first end cap is sized to fit entirely within saidcartridge.
 23. A chromatography column comprising: a column having aflexible-wall, said column containing chromatography media, an end capwithin said column, said end cap defining an inlet passage for flow ofprocess fluid and flow distributor passages for distributing processfluid across a cross-sectional area of said column, and a clamp locatedaround an outside of said column for fixing said end cap within saidcolumn, said clamp including a conical member.
 24. The chromatographycolumn of claim 23, wherein said clamp radially compresses said flexiblewall to seal said end cap within said column.
 25. The chromatographycolumn of claim 23 wherein said end cap is sized to fit entirely withinsaid column.